Epo and its receptor (EpoR) are prime regulators of red cell production. EpoR function depends upon an upstream Jak2 kinase, which acts in part by phosphorylating conserved EpoR cytoplasmic sites for transduction factor binding. Unexpectedly, PY-null EpoR forms in transgenic and knock-in mice retain significant activity, with only apparently mild defects observed in vivo. In new work, we have adopted an EpoR knock-in system, and in PY-null "EpoR-HM" mice have discovered: i) clear Epo dose-dependent defects in erythroid progenitor cell growth and development; ii) compensatory increases in serum growth factors; and iii) ineffective stress erythropoiesis. By comparison, the adding-back of a PY343 Stat5 binding site confers hyper-activity (EpoR-H form). Aim 1 will employ (anti)apoptotic factor analyses, RTqPCR, and global profiling to define core response pathways for EpoR-HM, EpoR-H and wt-EpoR receptors in purified primary cells. Roles for EpoR PY-dependent pathways in bcl-x gene induction will be investigated. For select factors, RNAi approaches will be used to assess functional significance. Aim 2 will examine stress erythropoiesis in EpoR-HM mice (including cisplatin and Cs137-induced anemia). EpoR-H mice also will be analyzed, as will underlying EpoR, Kit and/or glucocorticoid receptor linked mechanisms. Aim 3 will assess compensatory mechanisms invoked in EpoR-HM mice, including Epo production. The hypothesis that EpoR-H (and possibly EpoR-HM) function is assisted by a collateral (hematopoietic) growth factor-receptor system also will be investigated. Lyn kinase also appears to play important roles in EpoR action: Aim #4 will test the extent to which activity of the minimal EpoR form EpoR-HM depends upon Lyn by studying compound lyn -/-, EpoR-HM mice and assessing (advanced) erythropoietic defects. Such defects in lyn -/- mice per se also will be investigated. Aim #5 will use primary ceils from EpoR-H and EpoR-HM mice to extend new findings from cell line models that EpoR-PY343 and Kit PY567 support EpoR and Kit cofunction. This work in primary models should advance a basic understanding of core EpoR action mechanisms (including those essential for stress erythropoiesis). Insight also will be gained into the erythropoietic actions of Lyn and Kit PTK proto-oncogenes.